Lactobacilli species diversity in gut microbiota of renal failure patients

1 Introduction

The gastrointestinal tract of human consists of various bacteria including aerobic and anaerobic bacteria. The entire of colon is filled by mostly commensal and obligatory anaerobic bacteria such as lactobacilli (Muyzer et al., 1993; Dahroud et al., 2016; Quagliariello et al., 2016). Lactobacillus genus is a gram-positive, non-sporulating, catalase negative and rod-shaped bacillis that contains more than ninety validly described species and subspecies (Jabbari et al., 2017). Lactobacillus species are included in the division of Firmicutes (Ludwig et al., 2010) and are belonged to the members of lactic acid bacteria, which are found in human and animals intestinal microbiota (Vaughan et al., 2005; Ludwig et al., 2010). As well as, Lactobacillus species are one of the most commonly used probiotic strains in the wide range of food components in preventing chronic inflammation disease and enhancing health (Pandey et al., 2015). The fermentative characteristics of lactobacillus spp. have classified them into three divisions including obligate heterofermentative lactobacilli (OHEL), obligate homofermentative lactobacilli (OHOL) and facultative heterofermentative lactobacilli (FHEL)[24]. The activity of these aerobic and anaerobic bacteria has a major impact on the host health and characteristics such as development of the immune system, host protection against pathogens, and positive effects on the host nutrition and colonic health (Falk et al., 1998; Hooper et al., 2001; Gholizadeh et al., 2019). The protein-bound uremic toxins such as p-cresyl sulphate (PCS), indoxyl sulphate (IS), p-cresylglucuronide, kynurenic acid and indole acetic acid (IAA), which are normally excreted into the urine, could be contributing factor to progression chronic renal disease that have deleterious effects (Vanholder and Glorieux, 2015). These uremic toxins could lead to damage endothelial cells or renal tubular cells by fibrotic changes, inflammatory responses or enhancing cellular oxidative by activation of Nicotinamide adenine dinucleotide (NADPH) oxidase, P53, nuclear factor-ĸB (NFĸB) (Shimizu et al., 2013; Watanabe et al., 2013; Vanholder et al., 2014; Vanholder and Glorieux, 2015). The precursors of these uremic toxins are produced by gut microbiota from tyrosine or tryptophan and absorbed from the colon. Alters in the absorption of the uremic toxins from the colon may affect renal function (Vanholder and Glorieux, 2015) (Fig. 1).

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Fig. 1

The heat map generated from abundance mean of various species.

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Recent studies suggested that two third of individuals with abnormalities and a disequilibrium in the gastrointestinal and ecosystem have uremia (Kang, 1993). The majority of these abnormalities and disequilibrium happen in the colon and ileum that intestinal microbiota plays critical roles. The increase in aerobic bacteria such as Escherichia coli results in a decreasing in the anaerobic bacteria such as bifidobacteria and lactobacilli, as well as, generating toxic substrates, called uremic toxins (Simenhoff and Dunn, 1996; Vaziri et al., 2015). Several studies demonstrated that there is a deteriorated intestinal barrier in individuals with uremia, which is due to the abnormalities and disequilibrium of gut microbiota caused by the increase of pathogenic bacteria (Lenoir-Wijnkoop et al., 2007; Nakabayashi et al., 2010; Vaziri, 2012). Higher concentrations of urea and, consequently, increased ammonium are demonstrated in chronic renal diseases, which resulted in an increase pH that could promotes aerobic bacteria in the gastrointestinal and, subsequently, produce uremic toxins. Conversely, carbohydrates fermentation and acetic and lactic acid production by bifidobacteria and lactobacilli acidify the intestinal that could inhibit aerobic bacteria and normalize the intestinal microbiota in patients with chronic renal failure. In uremic individuals the fecal urease activity is increased at high concentrations of plasma urea. Therefore, a beneficial and consideration factor in uremic patients is the increase in colon microbial urease (Chow et al., 2003). Chronic renal disease is a health problem that can lead to end-stage kidney disease (ESKD), which is required kidney transplantation therapy. The modulation of interrelationship between the gut and kidney may be a potential role to slow progression of chronic renal disease (Ritz, 2011). Several studies suggested that alters in the gut microbiota affects the gut environment, which could correlate with kidney disease (Chow et al., 2003; Anders et al., 2013; Ramezani and Raj, 2014). Rossi et al. (2012) demonstrated that the pre- and probiotic therapy could significantly reduce IS and PCS in the chronic renal disease population. In addition, Yoshifuji et al. (2015) demonstrated that Lactobacillus spp. supplementation are improved intestinal changes and renal damage through activation of Lactobacillus recognized TLR (toll-like receptor). In another study, Vaziri et al. (2013) demonstrated that the abundance of Lactobacillaceae and Prevotellaceae in the intestinal microbiota flora are significantly decreased in the uremic animals compared to the control animals. Therefore, the aim of this study is assessment of abundance of lactobacillus spp. in fecal flora of ESKD and kidney transplanted patients.

2 Methods and materials

3 Results

The characteristics of participants between control group did not differ with disease group in age (59.3 ± 7.89 vs. 53.20 ± 12.03), and gender (male: 10, female: 10 vs. male: 14 vs. female: 6). Total 651 strains from 40 fecal samples were identified, which 37 (5.68%) strains were identified as order Lactobacillales. The strains were classified into six family including Lactobacillaceae (18/37), Streptococcaceae (14/37), Enterococcaceae (1/37), Aerococcaceae (1/37), Carnobacteriaceae (1/370 and Leuconostocaceae (2/37). In addition, the strains were classified into nine genus and 37 species including Lactobacillus (17/37), Straptococcus (10/37), Lactococcus (4/37), Abiotrophia (1/37), Granulicatella (1/37), Enterotoccoccus (1/37), Weisella (1/37), Leuconostoc (1/37), and Pediococcus (1/37). The most abundance genus in both groups are Streptococcus (4.47% ± 3.67% in control group and 3.12% ± 2.47% in disease group) and Lactobacillus (3.41% ± 4.15% in control group and 1.78% ± 2.88% in disease group) and the lowest abundance genus in both groups are Pediococcus (0.00005% ± 0.00009% in control group and 0.0006% ± 0.0009% in disease group) and Leuconostoc (0.004% ± 0.003% in control group and 0.002% ± 0.001% in disease group). S. dentisiani (2711.9 ± 4067.4) and L. crispatus (2289.65 ± 6280) were the most abundant species in control group and P. parvulus (0.15 ± 0.63) and L. graminis (2.45 ± 5.44) were the lowest species. In disease group, S. salivarius subsp. salivarius (6436.2 ± 9572.81) and E. durans (1756.85 ± 3674.53) were the most abundant species and L. secaliphilus (1 ± 3.54) and S. hongkongensis (1.65 ± 4.47) were the lowest abundant. The abundance of various species are shown in Table 1. S. salivarius subsp. salivarius, S. dentisiani, G. adiacens, and L. oligofermentans were found in all control group. As well as, S. salivarius subsp. salivarius, E. durans and S. dentisiani were found in all disease group. Comparing the abundance mean of the strains revealed that there is no significant association between control group and disease group (P-value > 0.05), but the abundance mean of the strains in control group increase in compared to disease group (719.9 ± 1534 vs. 462.8 ± 1115, P = 0.275). In addition, the presence of order Lactobacillales in the individuals of control group was not significant difference in compared to the individuals of disease group (10.49 ± 1.071 vs. 10.27 ± 1.044, P = 0.848).

4 Discussion

This study describes the abundance and diversity of order Lactobacillales in two groups including control group and disease group described as ESKD and kidney transplanted patients. In control group, the abundance of Lactobacillales were lower than disease group, and their species composition was different in comprised to disease group.

Marteau et al. (2001) demonstrated that the rRNA of Lactobacillus-Enterococcus group in healthy human is 6.6% of fecal bacterial rRNA, which is more than our study that found 3.8% in control group and 2.38% in disease group. In addition, Stsepetova et al. detected lactobacilli in all fecal samples by real-time PCR and 90% by the culture method in the samples, and we found lactobacilli in all fecal samples, which could suggested that the NGS method is the best for assessment of fecal bacteria.

This study is one of the few research concerning abundance of lactobacilli and their effects on chronic kidney disease patients, specifically comparing in the two groups as control and chronic kidney disease patients groups. However, the importance of this type of study in kidney disease patients lies in the benefits that could be obtained if the decrease of lactobacilli promoted symptoms in chronic kidney disease patients.

It was reported that gut microbiota may be a key role that maintains the gut in patients with chronic kidney disease (Kotanko et al., 2006). In addition, progression of chronic kidney disease may promoted by uremic toxins, which are made by certain gut microbiota (Satoh et al., 2003). Several studies suggested that early probiotic therapy to remove various uremic toxins could be delayed the onset of ESKD in individuals with progressive chronic kidney disease. Based on the previous study by Alatriste et al. (2014), Yoshifuji et al. (2015) and Vaziri et al. (2013), a decreased of lactobacilli abundance are shown in chronic kidney disease. In addition, the population of butyrate-producing enzymes bacterial families such as Lactobacillaceae and Prevotellaceae are lower in patients with uremia, whereas the population of indole- or p-cresol-producing bacterial families such as Enterobacteriaceae, Clostridiaceae, and Verrocomicrobiaceae are higher compared to health individuals (Wong et al., 2014). Similar to previous study, Lactobacillaceae is lower in patients group compared to healthy individuals in the present study. Ranganathan et al. suggested that probiotics with combination of S. thermophiles, B. longum and L. acidophilus are significantly reduced BUN and enhanced well-being in patients with chronic kidney disease (Ranganathan et al., 2010). Whereas, we have not found L. acidophilus in both control and patients groups. While, L. fermentum, L. paracasei, L. kefiri, L. vaginalis, L. crispatus, L. salivarius and other families of order Lactobacillales were reduced two or more folds in patients group compared to control group. In addition, L. mucosa, L. taiwanensis and L. ruminis were increased two or more folds in patients group compared to control group. Therefore, it has been suggested that future studies are necessary to elucidate more precise mechanisms of different lactobacilli strains in chronic kidney disease, subsequently develop more efficient therapeutic strategies. Our data could provide relevant information for these future studies.

5 Conclusion

More diversity was exhibited in lactobacillus spp. in patients with chronic kidney disease compared to control group. Some of the species of Lactobacillaceae family such as L. acidophilus was not found in both patients and control groups. In addition, more of the species of Lactobacillaceae were decreased in patients group, whereas certain species were increased in patients group compared to control group. This diversity could suppose a novel mechanisms and therapeutic strategies to slow the progression of renal damage and delay the onset of ESKD in patients with chronic kidney disease.